Major Activities/Specific Objectives. The principal goal of our ongoing research effort is to develop an in depth mechanistic understanding of the MMP-independent activities of members of the TIMP family, in particular TIMP-2. We have identified the following specific objectives to obtain our goals: 1) examine the role of TIMPs in altering the growth and invasive potential of cancer cells in vitro; 2) study to effects of TIMPs on primary and metastatic tumor growth in vivo; 3) study the influence of TIMPs on recruitment of immune-modulatory cells (myeloid-derived suppressor cells (MDSC)) to the primary tumor and metastatic niche4) develop a better understanding of the uptake and role of intercellular TIMP. In prior experiments with forced expression of TIMP-2 in tumor cells we have observed suppression of primary tumor growth. The suppression of tumor growth was accompanied by a statistically significant decrease in tumor microvascular density count (CD 31+ or CD34+), a measure of antiangiogenic effects, as well as by increased tumor cell apoptosis (also possibly due to inhibition of angiogenesis). Somewhat unexpectedly, we also observed a decrease in focal adhesion kinase (FAK) in TIMP-2 expressing tumors and a significant decrease in FAK phosphorylation (Y397) in both TIMP-2 and Ala+TIMP-2 expressing tumor cells. Our observation that both FAK and/or AKT (Protein Kinase B, PKB) phosphorylation is reduced in TIMP-2 and Ala+TIMP-2 tumor tissues is significant in that: 1) FAK is upstream of AKT signaling, and both are involved in regulation of cell migration; 2) TIMP-2 and Ala+TIMP-2 expression reduced tumor cell migration in vitro. We previously reported decreased FAK phosphorylation in endothelial cells where it is involved in control of eNOS activity. A major focus in my lab has been to examine the effects of exogenous TIMP-2 in murine tumor models. We are studying the effects on tumor growth and metastasis with the aim of developing a better understanding of the mechanisms that may affect these processes.These recent findings suggest that TIMP-2 has a variety of effects on both tumor and host cells that combine to produce a potent anti-tumor activity that could be exploited clinically.